The proposed research is designed to investigate certain of the biochemical and physiological processes that are fundamental to the maintenance of a normally functional cornea. In order that the tissue be transparent a constant level of hydration must be maintained in the stroma and this necessitates a balance between imbibition of fluid by the stroma and its extrusion across the endothelium. This equilibrium is determined by the permeability of the endothelial cell layer to water and solutes and by the efficiency of the energy-dependent process of active ion transport. Since lipids are critical for the maintenance of both normal permeability characteristics of cell membranes and normal activity of lipoprotein enzyme complexes, it is possible that failure of corneal homeostasis may result from degeneration of these compounds. It is postulated that under normal, in vivo, circumstances the cornea is subjected to a continuous attack by oxidants, such as H2O2 in the aqueous humor and free radicals generated by oxygen reactions in the cell, and that these are rendered innocuous by anti-oxidants, detoxifying enzymes and protective enzyme systems. In vitro, or under conditions of added stress that might be due to pathologies, aging, or excessive exposure to light, these protective mechanisms may be over-extended, allowing peroxidation of lipids and free radical damage to proteins and carbohydrates, with ensuing physiological failure. This hypothesis will be tested by measuring metabolic reactions, enzyme activities and redox state and concentration of key metabolites following exposure of corneas to H2O2 or UV light. Other adverse conditions will be tested for cumulative effects of stress, while added anti-oxidants or enzymes will be assessed for protective capacities. The status of these systems and their vulnerability to oxidative challenge will also be measured in diabetic animals to determine whether corneal problems encountered in diabetic vitrectomy patients might be due to their failure. The mechanisms of the fluid pump and its response to conditions described above will also be examined. Ion fluxes across the endothelium and cation and anion exchange process will be measured in order to determine the primary ion(s) transported and the role of protons, CO2 and carbonic anhydrase in fluid movement.